Helminths, known as ‘masters of immunomodulation’, use several immunomodulatory strategies to evade and/or modify immune responses in order to survive into the mammalian host for long periods of time. The parasitic helminth Fasciola hepatica causes fascioliasis, an emerging important human disease that affects around 17 million persons worldwide. Fascioliasis also infects livestock, causing economic losses estimated at more than $3 billion annually. As with many other helminths, F. hepatica polarizes the immune system of the host to a dominant Th2/T-regulatory status with suppression of inflammatory responses. As a result, hosts infected with F. hepatica are rendered more susceptible to secondary bystander infections, such as with Bordetella pertussis and Mycobacterium tuberculosis, which require Th1 immunity for protection.
The potent immune suppression exerted by F. hepatica is mediated by the copious amounts of excretory-secretory products (ESPs) released by the parasite, particularly the cathepsin-L peptidases, which represent approximately 80% of ESPs. Studies have demonstrated that ESPs of F. hepatica can mimic the immunomodulatory effect that is observed during active infection, without the tissue pathology, and also suppress the development of the Th1 response. For example, administration of F. hepatica Cathepsin-L1 (CatL1) cysteine protease suppressed the onset of protective Th1 immune responses to bacterial infections in mice and prevented the development of a Th1 response in mice inoculated with B. pertussis vaccine. Glutathione S-transferase (GST), another major antigen comprising 4% of ESPs, inhibited the proliferation of rat spleen cells in response to ConA stimulation in vitro. Both CatL1 and GST were shown to partially activate dendritic cells (DCs) via toll-like receptor-4 (TLR4), a pattern recognition receptor (PRR), using different intracellular signaling pathways. Other F. hepatica polypeptides that also play a role in host immunomodulation are the tegument antigens. The tegument constitutes the parasite-host interface and is the place where much of the immune interplay between the fluke and host occur. F. hepatica tegument antigens have been shown to significantly suppress the serum levels of gamma interferon (IFNγ) and interleukin-12p′70 (IL12p70) and to suppress expression of the cell-surface markers CD80, CD86 and CD40 by targeting multiple TLRs of DCs. Moreover, F. hepatica tegument antigens have also been shown to impair DC function in a mouse model of septic shock by inhibiting their phagocytic capacity and ability to prime T cells.
Proteomic studies have demonstrated that both ESPs and tegument antigens consist of highly complex mixtures of polypeptides that include proteolytic enzymes, transporters, membrane-associated proteins, antioxidants and many other trematode-specific proteins. Members of the 12 kDa fatty acid binding protein (FABP) family have been identified in most of these studies. FABPs play an essential role in parasite nutrition and have been recently categorized as antioxidant molecules. These proteins can potentially prevent oxidative damage to trematode cellular components by binding fatty acids and ions involved in oxidative stress. Previous studies have shown that vaccines containing FABPs induce partial immune protection in experimentally infected mice and sheep. Moreover, F. hepatica FABPs also appear to be important molecules for inducing cross-immunity against Schistosoma species. Although numerous published papers have explored the vaccine potential of F. hepatica FABPs, no studies have investigated whether FABPs have anti-inflammatory effects or whether they may interact with cells of the host immune system.
In the present invention, we purified native forms of FABP (named Fh12) from adult fluke extract for the anti-inflammatory properties of the purified protein in vitro and in vivo. This invention is the first to present the anti-inflammatory properties of F. hepatica fatty acid binding protein, providing evidence of its mechanism of action.